Biology subjects

Kamuf-Schenk, V.

Publications and source records attributed to Kamuf-Schenk, V..

6 recordsLinked to original sources

bioRxiv2026

Muscle LIM Protein is a Non-Canonical RNA-Binding Protein that Engages in RNA-Dependent Cytoskeletal Assemblies

Muscle LIM Protein (MLP) is a key component of the cardiomyocyte Z disc that is essential for sarcomere integrity and cardiac homeostasis. Accordingly, loss of MLP function results in (cardio)myopathy phenotypes in animal models and human patients. In this study, we present the first RNA-binding proteome (RBPome) of human cardiomyocytes using enhanced RNA interactome capture (eRIC). Data integration with existing RBPome datasets identified MLP as an evolutionary conserved and previously unrecognized non-canonical RNA-binding protein (RBP) in cardiomyocytes. Using complementary biochemical approaches, we confirmed direct RNA association of MLP. Moreover, RNA integrity was required for MLP interactions with cytoskeletal proteins and for the formation of higher-order MLP-containing assemblies, revealed by co-immunoprecipitation and RNA-dependent sedimentation profiling. Subcellular fractionation and imaging analyses indicated that MLPs association with cytoskeletal complexes may be weaker and more dynamic compared to core sarcomere proteins like -actinin. Domain mapping identified two glycine-rich regions within MLP as RNA contact sites, and RNA-binding-deficient MLP mutants showed impaired association with cytoskeletal complexes. Together, our findings suggest RNA binding as a regulated property of MLP and uncover an RNA-dependent layer of cytoskeletal organization in cardiomyocytes.

molecular biology↗

bioRxiv2026

Comprehensive analysis of nonsense-mediated mRNA decay targets and activity in cardiomyocytes

Nonsense-mediated mRNA decay (NMD) serves as a mechanism to suppress the expression of mutant alleles containing premature termination codons, limit the expression of aberrantly spliced transcript isoforms, and control the expression of numerous regular genes. While the principles by which NMD recognizes target transcripts are well understood, much less is known about the range of mRNAs subject to NMD in tissues and specialized cell types. Here we describe the landscape of genes whose expression is controlled by NMD in cardiomyocytes derived from human induced pluripotent stem cells (iPSC-CM), using small read RNA sequencing in combination with a potent inhibitor of SMG1, a kinase essential for NMD. We find that NMD targets are highly conserved between iPSC-CM lines derived from two healthy individuals. Beyond gene level analysis, we identify individual exon and intron RNA sequences that strongly accumulate upon SMG1 inhibition. Using the cardiac NMD targets identified at gene, exon and intron level, we then demonstrate reduced NMD efficiency upon knockdown of two essential NMD factors, UPF1 and UPF2, in iPSC-CM. Our analysis demonstrates that quantifying the transcriptome-wide response to SMG1 inhibition represents a highly sensitive approach to assess global activity of the NMD pathway.

molecular biology↗

bioRxiv2025

ZEB1 Is a Key Regulator of Cardiomyocyte Structure and Function

BackgroundOur previous work identified the transcription factor "Zinc Finger E-Box Binding Homeobox 1" (ZEB1) as a downstream effector of Cytoplasmic Polyadenylation Element Binding Protein 4 (CPEB4), an RNA-binding protein responsive to cardiac stress. While ZEB1 is known for its role in cancer metastasis and epithelial-to-mesenchymal transition (EMT), its function in cardiomyocytes is not well understood. Based on previous findings, we hypothesize that ZEB1 is essential for maintaining the structural integrity and mitochondrial function of cardiomyocytes. MethodsAAV9-Zeb1 was used for the overexpression of Zeb1. Using a myosin heavy chain alpha (MHC) Cre system, we created a Zeb1 conditional knockout mouse. To evaluate cardiac function and structure, we used echocardiography, electron microscopy, immunohistochemistry. We identified differentially expressed genes following Zeb1 deletion using RNA-seq and determined direct Zeb1 target genes by integrating this transcriptome data with a Zeb1 ChIP-seq dataset. ResultsZEB1 deletion leads to sarcomere damage, mitochondrial dysfunction, and dedifferentiation, with more pronounced effects in females. Overexpression promotes hypertrophic remodeling. Echocardiographic analysis showed progressive systolic dysfunction, and histology revealed sarcomeric disarray, again especially in females. A tamoxifen-inducible ZEB1 knockout mouse model only confirmed ZEB1s crucial role in fully differentiated cardiomyocytes in female mice. Integrated analysis of RNA-seq and ChIP-seq revealed that Zeb1 directly regulated mitochondrial genes, thereby playing a critical role cardiomyocyte energy supply and having secondary effects on cardiac structure and function. ConclusionsZEB1 is critical for cardiomyocyte homeostasis, and maintaining its function is necessary for normal cardiac performance and structure.

molecular biology↗

bioRxiv2022

Ythdf2 regulates cardiac remodeling through its m6A-mRNA target transcripts

m6A mRNA methylation controls cardiomyocyte function and increased overall m6A levels are a stereotyping finding in heart failure independent of the underlying etiology. However, it is largely unknown how the information is read by m6A reader proteins in heart failure. Here we show that the m6A reader protein Ythdf2 controls cardiac function and identified a novel mechanism how reader proteins control gene expression and cardiac function. Deletion of Ythdf2 in cardiomyocytes in vivo leads to cardiac hypertrophy, reduced heart function, and increased fibrosis during pressure overload as well as during aging. Similarly, in vitro the knockdown of Ythdf2 results in cardiomyocyte growth and remodeling. Mechanistically, we identified the eucaryotic elongation factor 2 as a major target of Ythdf2 using cell type specific Ribo-seq data. Our study expands our understanding on the regulatory functions of m6A methylation in cardiomyocytes and how cardiac function is controlled by the m6A reader protein Ythdf2.

molecular biology↗

bioRxiv2022

Transient inhibition of translation improves long-term cardiac function after ischemia/reperfusion by attenuating the inflammatory response

RationaleRapid reperfusion is the most effective treatment for attenuating cardiac injury caused by myocardial ischemia. Yet, reperfusion itself elicits damage to the myocardium through incompletely understood mechanisms, known as ischemia/reperfusion (I/R) injury. The myocardium adapts to I/R by changes in gene expression, which determines the cellular response to reperfusion. Protein translation is a key component of gene expression. However, it is unknown how regulation of translation contributes to cardiac gene expression in response to reperfusion and whether it can be targeted to mitigate I/R injury. MethodsTo examine translation and its impact on gene expression in response to I/R we assessed protein synthesis at different timepoints after ischemia and reperfusion in vitro and in vivo. Pharmacological inhibitors were used to dissect the underlying molecular mechanisms of translational control. Transient inhibition of protein synthesis was undertaken to decipher the effects of the translational response to reperfusion on cardiac function and inflammation. Cell-type-specific ribosome profiling was performed in mice subjected to I/R to determine the impact of translation on the regulation of gene expression in cardiomyocytes. ResultsReperfusion increased translation rates from a previously suppressed state during ischemia in cardiomyocytes, which was associated with the induction of cell death. In vivo, I/R resulted in strong activation of translation in the myocardial border zone. Detailed analysis revealed that the upregulation of translation is mediated by eIF4F complex formation, which was specifically mediated by the mTORC1-4EBP1-eIF4F axis. Short-term pharmacological inhibition of eIF4F complex formation by 4EGI-1 or rapamycin, respectively, attenuated translation, reduced infarct size and improved long-term cardiac function after myocardial infarction. Cardiomyocyte-specific ribosome profiling identified that reperfusion damage increased translation of mRNA networks in cardiomyocytes associated with cardiac inflammation and cell infiltration. Transient inhibition of the mTORC1-4EBP1-eIF4F axis decreased the expression of proinflammatory transcripts such as Ccl2, thereby reducing Ly6Chi monocyte infiltration and myocardial inflammation. ConclusionsMyocardial reperfusion induces protein synthesis in the border zone which contributes to I/R injury by rapidly translating a specific maladaptive mRNA network that mediates immune cell infiltration and inflammation. Transient inhibition of the mTORC1-4EBP1-eIF4F signaling axis during reperfusion attenuates this proinflammatory translational response, protects against I/R injury and improves long-term cardiac function after myocardial infarction. Clinical PerspectiveO_ST_ABSWhat Is New?C_ST_ABSO_LIThis is the first study to investigate the impact of translational regulation on cardiomyocyte gene expression in response to myocardial ischemia/reperfusion. C_LIO_LIWe show that translation regulates approximately two-thirds of differentially expressed genes in cardiomyocytes after ischemia/reperfusion, including many involved in inflammation and immune cell infiltration. C_LIO_LIThe translational response to ischemia/reperfusion is regulated by the mTORC1-4EBP1-eIF4F axis, which determines pro-inflammatory monocyte infiltration via control of the expression of the chemokine Ccl2. C_LI What Are the Clinical Implications?O_LICurrently, there are no specific therapies to prevent ischemia/reperfusion injury, which is mediated, at least in part, by a maladaptive inflammatory response. C_LIO_LIA translationally controlled network regulated by the mTORC1-4EBP1-eIF4F axis can be targeted by a short-term pharmacological intervention to attenuate the inflammatory response and improve cardiac function after ischemia/reperfusion in mice. C_LIO_LIThis study supports the emerging concept of selectively inhibiting maladaptive elements of the inflammatory response to improve outcome in patients after myocardial infarction; in addition, it provides a mechanistic basis for the currently ongoing CLEVER-ACS trial. C_LI

molecular biology↗

bioRxiv2020

Muscle specific translational control of Cand2 by mTORC1 regulates adverse cardiac remodeling

The mechanistic target of rapamycin (mTOR) is a key regulator of pathological remodeling in the heart by activating ribosomal biogenesis and mRNA translation. Inhibition of mTOR in cardiomyocytes is protective, however, a detailed role of mTOR in translational regulation of specific mRNA networks in the diseased heart is largely unknown. A cardiomyocyte genome-wide sequencing approach was used to define mTOR-dependent post-transcriptional gene expression control at the level of mRNA translation. This approach identified the muscle-specific protein Cullin-associated NEDD8-dissociated protein 2 (Cand2) as a translationally upregulated gene, dependent on the activity of mTOR. Deletion of Cand2 protects the myocardium against pathological remodeling. Mechanistically, we found that Cand2 links mTOR signaling to pathological cell growth by increasing Grk5 protein expression. Our data suggest that cell-type-specific targeting of mTOR might have therapeutic value for adverse pathological cardiac remodeling.

molecular biology↗